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Wizard miniprep protocol

Steve Stelman steves at burn.ucsd.edu
Tue Dec 12 20:15:59 EST 1995


Surasakdi Wongratanacheewin wrote:

> Does anyone there have the protocol for Wizard miniprep (Promega).
> I have the kit but I loss my protocol. Could you please send me that
> protocol.
> 
>         Thanks for help.
> 
> Surasak


        1) Pellet 1-3 ml cells.
           Resuspend in 200 ul Cell Resuspension Soln. (I)
           Add 200 ul Cell Lysis Soln. (II)
           Invert until soln clears.
           Add 200 ul Neutralization Soln. (III)
           Invert.
           Spin.
           Decant sup.
        
        2) Add 1 ml Purification Resin.
           Invert.
           Attach 5 ml syringe to column.  Remove plunger.
           Load sample into syringe.
           Gently push into column.
           Detach column, remove plunger, replace column.
        
        3) Load 2 ml Column Wash Soln.
           Gently push into column.
           Remove column to epitube.
           Spin dry 2 min.
        
        4) Remove column to clean epitube.
           Load 50 ul TE buffer.
           Wait 1 min.
           Spin 2 min. to elute.
        __________________________________________________
        (I) Cell Resuspension Soln:
                   50 mM  Tris-HCl, pH 7.5
                   10 mM  EDTA
                100ug/ml  RNase A

        (II) Cell Lysis Soln:
                  0.2 M  NaOH
                     1%  SDS

        (III) Neutralization Soln:
                 2.55 M  KOAc, pH4.8

        Column Wash Soln:
                  200mM  NaCl
                   20mM  Tris-HCl, pH 7.5
                    5mM  EDTA
               **dilute 1:1 with 95% EtOH  

PS.  If you want the recipe for the resin (or a working facsimile),
I have a recipe from an old post to this group for "Merlin" resin.



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